摘要
背景:以往研究发现修复神经损伤的细胞来源主要有许旺细胞、嗅鞘细胞、神经干细胞、激活的巨噬细胞等,但这些细胞存在着来源困难、有成瘤性、异体排斥等缺点。而羊膜上皮细胞不存在以上缺陷,且具有多向分化潜能,经诱导后可向心肌样细胞、神经干细胞、肝细胞、成骨和软骨细胞等分化。目的:体外培养大鼠羊膜上皮细胞,并诱导其向类神经干细胞方向分化。方法:取妊娠晚期大鼠,采用胰酶消化法获得羊膜上皮细胞,用无血清的神经干细胞条件培养基对细胞进行诱导分化,并用免疫细胞化学法和RT-PCR对诱导前后的细胞相关标志物进行鉴定。结果与结论:形态学观察结果显示,条件培养基诱导后的羊膜上皮细胞胞体回缩,胞核部分折光性增强,出现类似于树突及轴突样结构,这些突起可交织成网状,细胞贴壁牢靠。免疫细胞化学检测结果显示,与诱导前相比,条件培养基诱导后的羊膜上皮细胞中巢蛋白、胶质纤维酸性蛋白荧光强度较强,而特异性胚胎抗原4、波形蛋白荧光强度较弱。RT-PCR检测显示,与诱导前相比,条件培养基诱导后的羊膜上皮细胞的巢蛋白、胶质纤维酸性蛋白、微管相关蛋白mRNA的表达增强,而平滑肌22α、Sox-2及NanogmRNA的表达减弱。说明体外诱导的大鼠羊膜上皮细胞可成功分化为类神经干细胞。
BACKGROUND：Previous studies have found that Schwann cel s, olfactory ensheathing cel s, neural stem cel s and activated macrophages may repair nerve injuries, but these cel s have some deficiencies, such as source restrictions, tumorigenicity, and al ograft rejection. Amniotic epithelial cel s have no above-mentioned limits, and possess multiple-differentiation potential. After the induction, amniotic epithelial cel s may differentiate into cardiomyocyte-like cel s, neural stem cel s, liver cel s, osteoblasts and chondrocytes. OBJECTIVE：To culture rat amniotic epithelial cel s which can be induced to differentiate into neural-like stem cel s. METHODS：Rat amniotic epithelial cel s were col ected from late pregnancy rats using digestion method. Serum-free medium of nerve stem cel s was used to induce rat amniotic epithelial cel s into neural-like stem cel s. Normal and induced cel s were identified by immunocytochemistry and reverse transcription-PCR methods. RESULTS AND CONCLUSION：The morphological observation showed that amniotic epithelial cel s cultured in the conditioned medium presented with cel body retraction and part of nuclei showed refractive enhancement;dendrite-and axon-like structures appeared, and these processes were woven into a mesh and adhered firmly. Immunocytochemistry results showed that the induced amniotic epithelial cel s had a slightly stronger expression of Nestin and glial fibril ary acidic protein, while the pluripotent markers SSEA-4 and Vimentin expressions were weakened. Reverse transcription-PCR test showed that the induced amniotic epithelial cel s had a higher mRNA expression of Nestin, glial fibril ary acidic protein and microtubule-associated protein, while the mRNA expression of mesodermal smooth muscle cel marker smooth muscle-22αand pluripotent markers Sox-2 and Nanog declined. These findings suggest that cultured rat amniotic epithelial cel s can be induced to differentiate into neural-like stem cel s.
出处
《中国组织工程研究》
CAS
CSCD
2013年第19期3503-3507,共5页
Chinese Journal of Tissue Engineering Research
基金
国家重点基础研究发展计划(973计划
No.2005CB724302)~~
关键词
干细胞
干细胞培养与分化
羊膜上皮细胞
诱导
分化
类神经干细胞
巢蛋白
胶质纤维酸性蛋白
微管相关蛋白
973项目
干细胞图片文章
stem cells
stem cell culture and differentiation
amniotic epithelial cells
induction
differentiation
neural-like stem cells
Nestin
glial fibrillary acidic protein
microtubule-associated protein
National ＂973＂ Programof China
stem cell photographs-containing paper
